Selection of sunflower for resistance to cadmium accumulation

Abstract

Sunflower is an important economy crop in Ukraine, characterized by a dominant influence on the formation of the world edible oil market. According to EU standards, sunflower cadmium accumulation should not be more than 0.05 mg/kg. Facing the increasingly severe problem of improving the soil polluted by cadmium, cultivation of sunflower varieties with low cadmium accumulation can effectively reduce the cadmium content in seeds, fundamentally solving the problem of cadmium entering the body through the crop such as sunflower. Therefore, this study carried out the screening of sunflower varieties with low cadmium content. In addition, physiological and transcriptomic mechanisms for Cd accumulation in sunflower were studied. In this experiment, 104 sunflower varieties were used as test materials, and the biomass and cadmium content were used as screening indices of pot experiments, hydroponics, and field experiment to screen out sunflower varieties with the significant difference in cadmium accumulation and provide experimental materials for studying the different mechanism of cadmium accumulation. According to the results of a vegetation experiment using an analytical background with a cadmium concentration of 1.0 mg/kg, it was established that the range of values of the concentration indicator of this metal in the above ground part of plants varied from 0.5 to 2.6 mg/kg. According to the results of research, within the defined range, the collection was differentiated into 5 groups. High resistance to cadmium accumulation in the aboveground part of plants (0.5 to 0.9 mg/kg) was noted in 11 samples: JB231AC, S2K670, UE0100938, IPS\125, 64\3, Polaris CL, Reason, 56\3, Snk630, UE0100977, and UE0100018. The share of this group in the total collection was 10.6%. The group with a very low level of resistance (cadmium content is more than 2.11 mg/kg) included 7 selection samples such as PR63LL01, UE0100056, UE0100715, 62\3, UE0100052, JG3, and UE0100060. The share of this group in the collection was 6.7%. According to the results of the vegetation experiment with the analyzing background, the range of values of the cadmium concentration indicator in the aboveground phytomass of sunflower plants was determined, which was 0.5–2.26 mg/kg. The results of the experiment provided the possibility of preliminary differentiation of the sunflower collection with the selection of groups with different levels of resistance to cadmium accumulation. The main determining feature of the collection study in condition of a field experiment with a natural background of cadmium (0.21 mg/kg) was the resistance of plants to the accumulation of the metal in the seeds. According to the results of a 3-year experiment, the range of the cadmium content index varied from 0.23 to 0.43. The average value of the cadmium content index in seeds for the collection was 0.34+0.01 mg/kg. The value of the variation coefficient was 12.12%. Comparison of the vegetation and field experiment data of the cadmium content in seeds showed the similarity of the results regarding the differentiation of the collection by the level of resistance to the cadmium accumulation. 63.8% of selected samples analyzed by the method of indirect assessment corresponded to the indicators obtained in the field experiment. Therefore, the method of indirect evaluation provides high efficiency of negative selection against the trait and can be used for the preliminary selection of samples with a high level of resistance to cadmium accumulation in seeds. A group with a high level of resistance was also characterized by a high share of common (joint) hybrids of 76.1%. In all cases of group inconsistency, samples were transferred to the next (higher- or lower-ranked) group. This indicates that the reduced proportion of shared samples in the groups with good and satisfactory resistance levels of 40.6 and 42.8% can be explained by the mismatch of the boundaries of the selected groups. The high proportion of common hybrids (more than 2/3) in groups with low and unsatisfactory resistance indirectly indicates the universality of genetic mechanisms of cadmium accumulation in vegetative organs and sunflower seeds. On the contrary, the decrease in the share of common hybrids in groups with high and good levels of resistance indirectly indicates the difference in the control mechanisms of cadmium accumulation processes in vegetative organs and seeds. According to the results of the basic collection assessment in terms of the field experiment, the average values, and the range of variation of the main selectively controlled traits were determined. The average indicator value of the duration period “seedling-flowering” was 56.5 days with a range of variability from 50 to 66 days. The coefficient of variation was 6.81%. The average value, the range of variability, and the variation coefficient of base collection for plant productivity were 33.6 g, 11.70 - 63.30 g, and 20.75%, respectively. According to the results of the cluster analysis, morphological heterogeneity was established in the group of low cadmium samples, with the selection of three intragroup clusters. The first cluster contained late-ripening (the duration of the “seedling-flowering” phase was > 64 days), tall (> 193 cm), and low-yielding samples, such as UE0100114, UE01000114, UE0100977, and UE0100018. The second cluster included samples with an average and higher level of plant productivity for the collection (22.6 - 52.0 g/plant). A separate cluster was formed by the samples with specific characteristics. The heterogeneity of the group of low-cadmium samples implies the availability of different mechanisms of seed productivity formation and implementation of the trait of resistance to cadmium accumulation. On the contrary, in the sample structure of the group with “high cadmium accumulation”, the availability of unified transportation mechanism of biologically active cadmium from vegetative organs to the seeds was observed. The heterogeneity of the basic collection of selection samples and the difference in the mechanisms of ensuring a low level of cadmium accumulation in sunflower seeds was confirmed by the results of inter-varietal crossings. 73 samples of inter-varietal hybrids were obtained and tested in F1 and F2. The following result was established: when crossing “low cadmium” parental forms, only a fifth or 19% of them inherited the resistance trait to cadmium accumulation by heterosis type. The shares of hybrids that inherited the trait by the type of partial positive dominance and intermediate type for parental forms amounted to 9.5% per each. In most cases, 62% of inheritance trait occurred according to the type of depression and partial negative dominance. Depression and partial negative dominance were also the predominant types of inheritance in crossing variants in the group of “high cadmium” breeding samples and the intergroup crossing. The total share of these types of phenotypic inheritance was 85.7% in the case of “high cadmium” samples crossing and it was 66.7% in intergroup crossings. In this study, two sunflower cultivars of 62\3 (high Cd) and JB231AC (low Cd), were chosen to compare physiological and transcriptomic responses at different Cd concentrations (0, 25, 50, 100 μM). The results showed that JB231AC had better Cd tolerance than 62\3. The contents of H2O2 and MDA (malondialdehyde) in 62\3 were lower than that in JB231AC under Cd stress, but the activities of SOD (superoxide dismutase) and POD (peroxidase) in JB231AC were higher than in 62\3, which indicated that JB231AC had a strong ability to remove reactive oxygen species (ROS)-induced toxic substances. Many deferentially expressed ABC (ATP-binding cassette) and ZIP (Zn-regulated transporter, Iron-regulated transporter-like protein) genes indicated that the two gene families might play important roles in different levels of Cd accumulation in the two cultivars. One up-regulated NRAMP (Natural resistance-associated macrophage protein) gene was identified and had a higher expression level in 62\3. According to the results of experimental research, the average value of cadmium content in seeds for the experiment was 0.41 mg/kg with a range of variation from 0.05 to 2.24 mg/kg. The maximum value of the indicator was for hybrid 19/11 (a combination of the commercial hybrid Polar CL and the Sumy NAU line 64/3). The largest share of hybrids with the minimal cadmium content (higher or equal to 0.1) was obtained in combinations with line 56\3 and commercial hybrids Rezon and Sumico as the maternal component. The minimum content of cadmium was noted in combinations with the parent component (pollinator) of commercial hybrid Sumico with X51B and JB231AC lines. These results provide valuable information for further understand the mechanism of Cd accumulation and supply source materials for breeding of new high- or low- Cd sunflower cultivars.